Role of the UL25 protein in herpes simplex virus DNA encapsidation.
The herpes simplex virus protein UL25 attaches to the external vertices of herpes simplex virus type 1 capsids and is required for the stable packaging of viral DNA. To define regions of the protein important for viral replication and capsid attachment, the 580-amino-acid UL25 open reading frame was disrupted by transposon mutagenesis. The UL25 mutants were assayed for complementation of a UL25 deletion virus, and in vitro-synthesized protein was tested for binding to UL25-deficient capsids. Of the 11 mutants analyzed, 4 did not complement growth of the UL25 deletion mutant, and analysis of these and additional mutants in the capsid-binding assay demonstrated that UL25 amino acids 1 to 50 were sufficient for capsid binding. Several UL25 mutations were transferred into recombinant viruses to analyze the effect of the mutations on UL25 capsid binding and on DNA cleavage and packaging. Studies of these mutants demonstrated that amino acids 1 to 50 of UL25 are essential for its stable interaction with capsids and that the C terminus is essential for DNA packaging and the production of infectious virus through its interactions with other viral packaging or tegument proteins. Analysis of viral DNA cleavage demonstrated that in the absence of a functional UL25 protein, aberrant cleavage takes place at the unique short end of the viral genome, resulting in truncated viral genomes that are not retained in capsids. Based on these observations, we propose a model where UL25 is required for the formation of DNA-containing capsids by acting to stabilize capsids that contain full-length viral genomes.
Cockrell
Shelley K
SK
Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, W1256 Biomedical Science Tower, Pittsburgh, PA 15261, USA. flhoma@pitt.edu
Sanchez
Minerva E
ME
Erazo
Angela
A
Homa
Fred L
FL
D000818:Animals; D001665:Binding Sites; D002522:Chlorocebus aethiops; D004251:DNA Transposable Elements; D004279:DNA, Viral / Q000378:metabolism; D017353:Gene Deletion; D005816:Genetic Complementation Test; D018259:Herpesvirus 1, Human / Q000502:physiology*; D016254:Mutagenesis, Insertional; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D025941:Protein Interaction Mapping; D014709:Vero Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly
D004251:DNA Transposable Elements; D004279:DNA, Viral; D050505:Mutant Proteins; D014764:Viral Proteins; C069185:DNA cleavage and packaging proteins, Herpesvirus